title: LacR mutations are frequently observed in Streptococcus intermedius and are responsible for increased intermedilysin production and virulence.
abstract: Streptococcus intermedius secretes a human-specific cytolysin, intermedilysin (ILY), which is considered to be the major virulence factor of this pathogen. We screened for a repressor of ily expression by using random gene disruption in a low-ILY-producing strain (PC574). Three independent high-ILY-producing colonies that had plasmid insertions within a gene that has high homology to lacR were isolated. Validation of these observations was achieved through disruption of lacR in strain PC574 with an erythromycin cassette, which also led to higher hemolytic activity, increased transcription of ily, and higher cytotoxicity against HepG2 cells, compared to the parental strain. The direct binding of LacR within the ily promoter region was shown by a biotinylated DNA probe pulldown assay, and the amount of ILY secreted into the culture supernatant by PC574 cells was increased by adding lactose or galactose to the medium as a carbon source. Furthermore, we examined lacR nucleotide sequences and the hemolytic activity of 50 strains isolated from clinical infections and 7 strains isolated from dental plaque. Of the 50 strains isolated from infections, 13 showed high ILY production, 11 of these 13 strains had one or more point mutations and/or an insertion mutation in LacR, and almost all mutations were associated with a marked decline in LacR function. These results strongly suggest that mutation in lacR is required for the overproduction of ILY, which is associated with an increase in pathogenicity of S. intermedius.
authors.affiliation: Department of Biological Science and Technology, Institute of Technology and Science, The University of Tokushima Graduate School, Tokushima, Japan.
authors.forename: Toshifumi
authors.initials: T
authors.lastname: Tomoyasu
authors.forename: Hidenori
authors.initials: H
authors.lastname: Imaki
authors.forename: Sachiko
authors.initials: S
authors.lastname: Masuda
authors.forename: Ayumi
authors.initials: A
authors.lastname: Okamoto
authors.forename: Hyejin
authors.initials: H
authors.lastname: Kim
authors.forename: Richard D
authors.initials: RD
authors.lastname: Waite
authors.forename: Robert A
authors.initials: RA
authors.lastname: Whiley
authors.forename: Ken
authors.initials: K
authors.lastname: Kikuchi
authors.forename: Keiichi
authors.initials: K
authors.lastname: Hiramatsu
authors.forename: Atsushi
authors.initials: A
authors.lastname: Tabata
authors.forename: Hideaki
authors.initials: H
authors.lastname: Nagamune
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001430:Bacteriocins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D045744:Cell Line, Tumor; D056945:Hep G2 Cells; D006801:Humans; D056968:Lac Repressors / Q000235:genetics*; D055017:Monocyte-Macrophage Precursor Cells / Q000378:metabolism / Q000382:microbiology; D016254:Mutagenesis, Insertional; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D013290:Streptococcal Infections / Q000235:genetics / Q000378:metabolism / Q000382:microbiology*; D034367:Streptococcus intermedius / Q000235:genetics* / Q000378:metabolism / Q000472:pathogenicity; D014774:Virulence / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D001430:Bacteriocins; D056968:Lac Repressors; C403910:intermedilysin protein, Streptococcus intermedius